A fatal case of diphtheria diagnosed by antigen and gene detection, July 1999 - Gifu
(IASR 1999; 20: 302-303)

According to IASR Vol.19, No.10 (1998), only one fatal diphtheria case was reported in Japan last a decade. Thus, very few Japanese physician experience the disease, and it hampers correct clinical diagnosis.

A 68 year old male was transferred to a public hospital in Gifu Prefecture on July 31, 1999, because of progressive difficulty of breathing. Upon arrival, the patient presented symptoms suspecting suffocation, so that attending physician attempted intubation immediately. Because, white tumor-like lesion on the pharynx and larynx disturbed to maintain visual contact to the airway, intratracheal intubation was very hard. Soon after he developed heart arrest and respiratory arrest, and resuscitation was attempted but it was failed. A laryngologist examined after he died. The pharynx and larynx changed tumor-like white lesion. Because autopsy was not allowed, tissue on the lesion was collected.

Microscopic observation revealed acute inflammatory and highly edematous lesion with neutrophilic infiltration. Mucosal tissue was covered with thick pus, suspecting acute infectious laryngitis. Neutrophilic infiltration, bleeding, and remarkable fibrin infiltration was observed in the pus layer. Numerous Gram-positive rod was seen in the pus layer. Histological study strongly suspected diphtheria infection, because Gram-positive bacteria infection is rare on the lesion and acute reasonable clinical onset and death due to respiratory failure.

Bacterial identification was conducted using the pathological specimen and pus floated in formalin. Gram-positive bacterial clusters were detected from these specimens. Metachromatic body stain showed the few bacteria with variant colored club-like ends. Scanning electron microscopic observation revealed length of the bacterium was 1-2 um with clubbed ends, which was correspond with Corynebacterium diphtheriae. Anti-diphtheria fluorescent antibody test was conducted using paraffin-blocked specimen and formalin-fixed smear. Positive fluorescent was observed on the rod bacteria. Detection of diphtherial toxin from paraffin-blocked specimen was requested to the Department of Bacterial and Blood Products, NIID. Polymerase chain reaction (PCR) did not detect the gene for diphtheria toxin. Thus, specific PCR primer sets to detect shorter DNA fragments were prepared. Positive band, which is correspond with diphtheria toxin, was obtained. Its DNA sequence was analyzed and 60 base pair of partial gene for diphtheria toxin was identified.

These results concluded that this case is a diphtheria infection.

Reported by Masafumi Ito and Toshiaki Suzuki, Dept. of Pathology, Nagoya University Hospital.

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